LOCUS       sequence-205153-        4329 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Formaldehyde biosensor plasmid with sGFP reporter and modified frm 
            promoter.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4329)
  AUTHORS   Woolston BM, Roth T, Kohale I, Liu D, Stephanopoulos G
  TITLE     Development of a Formaldehyde Biosensor with Application to 
            Synthetic Methylotrophy.
  JOURNAL   Biotechnol Bioeng. 2017 Sep 16. doi: 10.1002/bit.26455.
  PUBMED    28921510
REFERENCE   2  (bases 1 to 4329)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Dec 15, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "Biotechnol 
            Bioeng. 2017 Sep 16. doi: 10.1002/bit.26455."
COMMENT     Sequence Label: pTR47m4-GFP
FEATURES             Location/Qualifiers
     source          1..4329
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      996..1218
                     /label=pSC101 ori
                     /note="low-copy replication origin that requires the Rep101
                     protein"
     CDS             join(1895..1897,1898..2611)
                     /codon_start=1
                     /product="GFP variant that folds robustly even when fused 
                     to poorly folded proteins (Pedelacq et al., 2006)"
                     /label=superfolder GFP
                     /note="mammalian codon-optimized"
                     /translation="VSKGEELFTGVVPILVELDGDVNGHKFSVRGEGEGDATNGKLTLK
                     FICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKRHDFFKSAMPEGYVQERTISFKDDG
                     TYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNFNSHNVYITADKQKNGIKA
                     NFKIRHNVEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSVLSKDPNEKRDHMVLLE
                     FVTAAGITHGMDELYK"
     promoter        3056..3110
                     /gene="tetR"
                     /label=tetR/tetA promoters
                     /note="overlapping promoters for bacterial tetR and tetA"
     protein_bind    3057..3075
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    3087..3105
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O1 for the tetR and tetA genes"
     protein_bind    3111..3129
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     CDS             complement(join(3255..4046,4047..4115))
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNKAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     3878..3897
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(4116..4220)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     4288..4306
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     CDS             complement(join(4327..4329,1..948))
                     /codon_start=1
                     /gene="rep101"
                     /product="RepA protein needed for replication with the 
                     pSC101 origin"
                     /label=Rep101
                     /translation="MSELVVFKANELAISRYDLTEHETKLILCCVALLNPTIENPTRKE
                     RTVSFTYNQYAQMMNISRENAYGVLAKATRELMTRTVEIRNPLVKGFEIFQWTNYAKFS
                     SEKLELVFSEEILPYLFQLKKFIKYNLEHVKSFENKYSMRIYEWLLKELTQKKTHKANI
                     EISLDEFKFMLMLENNYHEFKRLNQWVLKPISKDLNTYSNMKLVVDKRGRPTDTLIFQV
                     ELDRQMDLVTELENNQIKMNGDKIPTTITSDSYLRNGLRKTLHDALTAKIQLTSFEAKF
                     LSDMQSKHDLNGSFSWLTQKQRTTLENILAKYGRI"
ORIGIN
        1 gatccttccg tatttagcca gtatgttctc tagtgtggtt cgttgttttt gcgtgagcca
       61 tgagaacgaa ccattgagat catgcttact ttgcatgtca ctcaaaaatt ttgcctcaaa
      121 actggtgagc tgaatttttg cagttaaagc atcgtgtagt gtttttctta gtccgttacg
      181 taggtaggaa tctgatgtaa tggttgttgg tattttgtca ccattcattt ttatctggtt
      241 gttctcaagt tcagttacga gatccatttg tctatctagt tcaacttgga aaatcaacgt
      301 atcagtcggg cggcctcgct tatcaaccac caatttcata ttgctgtaag tgtttaaatc
      361 tttacttatt ggtttcaaaa cccattggtt aagcctttta aactcatggt agttattttc
      421 aagcattaac atgaacttaa attcatcaag gctaatctct atatttgcct tgtgagtttt
      481 cttttgtgtt agttctttta ataaccactc ataaatcctc atagagtatt tgttttcaaa
      541 agacttaaca tgttccagat tatattttat gaattttttt aactggaaaa gataaggcaa
      601 tatctcttca ctaaaaacta attctaattt ttcgcttgag aacttggcat agtttgtcca
      661 ctggaaaatc tcaaagcctt taaccaaagg attcctgatt tccacagttc tcgtcatcag
      721 ctctctggtt gctttagcta atacaccata agcattttcc ctactgatgt tcatcatctg
      781 agcgtattgg ttataagtga acgataccgt ccgttctttc cttgtagggt tttcaatcgt
      841 ggggttgagt agtgccacac agcataaaat tagcttggtt tcatgctccg ttaagtcata
      901 gcgactaatc gctagttcat ttgctttgaa aacaactaat tcagacatac atctcaattg
      961 gtctaggtaa ttttaatcac tataccaatt gagatgggct agtcaatgat aattactagt
     1021 ccttttcctt tgagttgtgg gtatctgtaa attctgctag acctttgctg gaaaacttgt
     1081 aaattctgct agaccctctg taaattccgc tagacctttg tgtgtttttt ttgtttatat
     1141 tcaagtggtt ataatttata gaataaagaa agaataaaaa aagataaaaa gaatagatcc
     1201 cagccctgtg tataactcac tactttagtc agttccgcag tattacaaaa ggatgtcgca
     1261 aacgctgttt gctcctctac aaaacagacc ttaaaaccct aaaggcttaa gtagcaccct
     1321 cgcaagctcg ggcaaatcgc tgaatattcc ttttgtctcc gaccatcagg cacctgagtc
     1381 gctgtctttt tcgtgacatt cagttcgctg cgctcacggc tctggcagtg aatgggggta
     1441 aatggcacta caggcgcctt ttatggattc atgcaaggaa actacccata atacaagaaa
     1501 agcccgtcac gggcttctca gggcgtttta tggcgggtct gctatgtggt gctatctgac
     1561 tttttgctgt tcagcagttc ctgccctctg attttccagt ctgaccactt cggattatcc
     1621 cgtgacaggt cattcagact ggctaatgca cccagtaagg cagcggtatc atcaacaggc
     1681 ttacccgtct tactgtcaag aggacatccg gtcaaataaa acgaaaggct cagtcgaaag
     1741 actgggcctt tcgttttgct gaggagactt agggacccta aattttattc tccagtgtta
     1801 tatactatag gggggtatgc attgacatat agcatacact cctatagtat attgcgtgca
     1861 gataacctgc aggtgcagta aggaggaaaa aaaagtgagc aagggcgaag agctttttac
     1921 cggtgttgtg ccgattttag tagaactgga cggagacgtg aacggtcata agttctctgt
     1981 tcgtggcgaa ggagagggag atgccaccaa tggtaagctg accctgaagt tcatctgtac
     2041 caccggtaag ctgcccgtgc cttggccgac gctggtcaca acgttgacgt atggcgtcca
     2101 atgcttttca cgctatccag atcacatgaa acgccacgac ttttttaaaa gcgcaatgcc
     2161 tgaaggttat gtgcaggaac ggactattag cttcaaagac gatgggacgt ataagacccg
     2221 cgcggaagtg aaatttgaag gcgatacctt agttaaccgc attgaattaa aaggtatcga
     2281 tttcaaagag gatgggaata tcctggggca caaattggaa tacaacttta attcgcacaa
     2341 cgtatacatt acagcggata aacagaaaaa tggcatcaaa gccaacttta aaatccgtca
     2401 taacgtagag gacggttccg tgcagctggc tgatcattac cagcagaata ctccgattgg
     2461 cgatggcccc gttctgctcc cggataatca ttacctgtct acacaaagcg ttcttagtaa
     2521 agacccaaac gagaagcgtg accatatggt cctgttggaa ttcgtcacgg cagcggggat
     2581 tactcatggc atggatgaac tctataagta aacggagcca atgtacgcaa aaaaccccgc
     2641 ttcagcgggg ttttttcgcc aaaaaaaacc ccgccctgtc aggggcgggg ttttttttta
     2701 gggccctact gactgtctat ttaagatagg ctcgaaccag ttcaatagtg tcgtcaacgg
     2761 attggctgac ttcgcggctg tagcagtcat ttcggtcaaa cgtttcccgg atatggcttt
     2821 caagcacttc tgccatcagc ccattagccg cgccccgaac ggcagcgatc tgttggagta
     2881 tggcacggca ttcggcatca ccctccagcg accgttccag agcatcaatc tgccctcgaa
     2941 tacgacgaac tcgagtaagg acctttttct tctcttccgg agtactgggc attttttttt
     3001 cctccttata agggggtagg gggtctagat ttttgtcgaa ctattcattt cacttttctc
     3061 tatcactgat agggagtggt aaaataactc tatcaatgat agagtgtcaa tctctatcac
     3121 tgatagggat catctggcca ttcgatggac ttaattaacg gcactgcgaa aaaaccccgc
     3181 cgaagcgggg ttttttgcga aatataatga ccctcttgat aacccaagag ggcatttttt
     3241 aatgcccatg gcgtttacca atgcttaatc agtgaggcac ctatctcagc gatctgtcta
     3301 tttcgttcat ccatagttgc ctgactcccc gtcgtgtaga taactacgat acgggagggc
     3361 ttaccatctg gccccagtgc tgcaatgata ccgcgagacc cacgctcacc ggctccagat
     3421 ttatcagcaa taaaccagcc agccggaagg gccgagcgca gaagtggtcc tgcaacttta
     3481 tccgcctcca tccagtctat taattgttgc cgggaagcta gagtaagtag ttcgccagtt
     3541 aatagtttgc gcaacgttgt tgccattgct acaggcatcg tggtgtcacg ctcgtcgttt
     3601 ggtatggctt tattcagctc cggttcccaa cgatcaaggc gagttacatg atcccccatg
     3661 ttgtgcaaaa aagcggttag ctccttcggt cctccgatcg ttgtcagaag taagttggcc
     3721 gcagtgttat cactcatggt tatggcagca ctgcataatt ctcttactgt catgccatcc
     3781 gtaagatgct tttctgtgac tggtgagtac tcaaccaagt cattctgaga atagtgtatg
     3841 cggcgaccga gttgctcttg cccagcgtca atacgggata ataccgcgcc acatagcaga
     3901 actttaaaag tgctcatcat tggaaaacgt tcttcggggc gaaaactctc aaggatctta
     3961 ccgctgttga gatccagttc gatgtaaccc actcgtgcac ccaactgatc ttcagcatct
     4021 tttactttca ccagcgtttc tgggtgagca aaaacaggaa ggcaaaatgc cgcaaaaaag
     4081 ggaataaggg cgacacggaa atgttgaata ctcatactct tcctttttca atattattga
     4141 agcatttatc agggttattg tctcatgagc ggatacatat ttgaatgtat ttagaaaaat
     4201 aaacaaatag gggttccgcg cacatttccc cgaaaagtgc cacctgacgt ctaagaaacc
     4261 attattatca tgacattaac ctataaaaat aggcgtatca cgaggccctt aggccaaata
     4321 ggccgttca
//
